The presence of an ER exit signal determines the protein sorting upon ER exit in yeast

In yeast, there are at least two vesicle populations upon ER (endoplasmic reticulum) exit, one containing Gap 1 p (general amino-acid permease) and a glycosylated alpha-factor, gp alpha F (glycosylated pro alpha-factor), and the other containing GPI (glycosylphospha-tidylinositol)-anchored proteins, Gas 1 p (glycophospholipid-anchored surface protein) and Yps 1 p. We attempted to identify sorting determinants for this protein sorting event in the ER. We found that mutant Gas 1 proteins that lack a GPI anchor and/or S/T region (serine- and threonine-rich region), two common characteristic features conserved among yeast GPI-anchored proteins, were still sorted away from Gap 1 p-containing vesicles. Furthermore, a mutant glycosylated alpha-factor, gp alpha GPI, which contains both the GPI anchor and S/T region from Gas 1 p, still entered Gap 1 p-containing vesicles, demonstrating that these conserved characteristics do not prevent proteins from entering Gap 1 p-containing vesicles. gp alpha F showed severly reduced budding efficiency in the absence of its ER exit receptor Erv29p, and this residual budding product no longer Gap 1 p-containing vesicles. These results suggest that the interaction of gp alpha F with Erv29p is essential for sorting into Gap 1 p-containing vesicles. We compared the detergent solubility of Gas 1 p and the gp alpha GPI in the ER with that in ER-derived vesicles. Both GPI-anchored proteins similarly partitioned into the DRM (detergent-resistant membrane) in the ER. Based on the fact that they entered different ER-derived vesicles, we conclude that DRM partitioning of GPI-anchored proteins is not the dominant determinant of protein sorting upon ER exit. Interestingly, upon incorporation into the ER-derived vesicles, gp alpha GPI was no longer detergent-insoluble, in contrast with the persistent detergent insolubility of Gas 1 p in the ER-derived vesicles. We present different explanations for the different behaviours of GPI-anchored proteins in distinct ER-derived vesicle populations.